This invention relates to relatively short peptides, and more particularly to peptides between about 10 and about 45 residues in length, which are naturally available in minute amounts in the venom of the cone snail or analogous to the naturally available peptides, and which include two or three cyclizing disulfide linkages. The peptides disclosed herein belong to a related family, the "A-lineage" conotoxin peptides.
The publications and other materials used herein to illuminate the background of the invention, and in particular, cases to provide additional details respecting the practice, are incorporated by reference, and for convenience are numerically referenced in the following text and respectively grouped in the appended bibliography.
Mollusks of the genus Conus produce a highly toxic venom which enables them to carry out their unique predatory lifestyle. Prey are immobilized by the venom which is injected by means of a highly specialized venom apparatus, a disposable hollow tooth which functions both in the manner of a harpoon and a hypodermic needle.
Few interactions between organisms are more striking than those between a venomous animal and its envenomated victim. Venom may be used as a primary weapon to capture prey or as a defense mechanism. These venoms disrupt essential organ systems in the envenomated animal, and many of these venoms contain molecules directed to receptors and ion channels of neuromuscular systems.
Predatory cone snails (Conus) have developed a unique biological strategy. Their venom contains relatively small peptides that are targeted to various neuromuscular receptors and may be equivalent in their pharmacological diversity to the alkaloids of plants or secondary metabolites of microorganisms. Many of these peptides are among the smallest nucleic acid-encoded translation products having defined conformations, and as such, they are somewhat unusual. Peptides in this size range normally equilibrate among many conformations. Proteins having a fixed conformation are generally much larger.
Cone snails that produce these toxic peptides, which are generally referred to as conotoxins or conotoxin peptides, are a large genus of venomous gastropods comprising approximately 500 species. All cone snail species are predators that inject venom to capture prey, and the spectrum of animals that the genus as a whole can envenomate is broad. A wide variety of hunting strategies are used; however, every Conus species uses fundamentally the same basic pattern of envenomation.
The major paralytic peptides in these fish-hunting cone venoms were the first to be identified and characterized. In C. geographus venom, three classes of disulfide-rich peptides were found: the .alpha.-conotoxin peptides (which target and block the nicotinic acetylcholine receptors); the .mu.-conotoxin peptides (which target and block the skeletal muscle Na.sup.+ channels); and the .omega.-conotoxin peptides (which target and block the presynaptic neuronal Ca.sup.2+ channels). However, there are multiple homologs in each toxin class; for example, there are at least five different .omega.-conotoxin peptides present in C. geographus venom alone. Considerable variation in sequence is evident, and when different .omega.-conotoxin peptide sequences were first compared, only the cysteine residues that are involved in disulfide bonding and one glycine residue were found to be invariant. Another class of conotoxins found in C. geographus venom is that referred to as conantokins, which cause sleep in young mice and hyperactivity in older mice and are targeted to the NMDA receptor. Each cone venom appears to have its own distinctive group, or signature, of different conotoxin sequences.
Many of these peptides have now become fairly standard research tools in neuroscience. .mu.-Conotoxin peptides, because of their ability to preferentially block muscle but not axonal Na.sup.+ channels, are convenient tools for immobilizing skeletal muscle without affecting axonal or synaptic events. .omega.-Conotoxin peptides have become standard pharmacological reagents for investigating voltage-sensitive Ca.sup.2+ channels and are used to block presynaptic termini and neurotransmitter release. The .alpha.-conotoxins have various clinical uses. One such use is their utility as clinical muscle relaxants because of their ability to achieve antagonistic blockage of the mammalian neuromuscular junction nAChRs. Another use for the .alpha.-conotoxins is in the diagnosis of myasthenia gravis (33, 34).
Some .alpha.-conotoxins have been found to bind preferentially to neuronal nicotinic receptors rather than to neuromuscular receptors. Such neuronal receptors are found on small-cell lung carcinomas (SCLCs). These .alpha.-conotoxins bind to the neuronal receptors and inhibit the proliferation of small-cell lung carcinomas which is caused by nicotine or cytosine stimulation.
Additional conotoxin peptides having the different general properties described above continue to be sought.